State of the art in hair analysis for detection of drug and alcohol abuse.

Prenatal substance abuse continues to be a significant problem in this country and poses important health risks for the developing fetus. The primary care pediatrician's role in addressing prenatal substance exposure includes prevention, identification of exposure, recognition of medical issues for the exposed newborn infant, protection of the infant, and follow-up of the exposed infant. This report will provide information for the most common drugs involved in prenatal exposure: nicotine, alcohol, marijuana, opiates, cocaine, and methamphetamine.

https://pediatrics.aappublications.org/content/pediatrics/early/2013/02/20/peds.2012-3931.full.pdf

Prenatal Substance Abuse: Short- and Long-term Effects on the Exposed Fetus

Drug testing, commonly used in health care, workplace, and criminal settings, has become widespread during the past decade. Urine drug screens have been the most common method for analysis because of ease of sampling. The simplicity of use and access to rapid results have increased demand for and use of immunoassays; however, these assays are not perfect. Falsepositive results of immunoassays can lead to serious medical or social consequences if results are not confirmed by secondary analysis, such as gas chromatography‐mass spectrometry. The Department of Health and Human Services’ guidelines for the workplace require testing for the following 5 substances: amphetamines, cannabinoids, cocaine, opiates, and phencyclidine. This article discusses potential false-positive results and false-negative results that occur with immunoassays of these substances and with alcohol, benzodiazepines, and tricyclic antidepressants. Other pitfalls, such as adulteration, substitution, and dilution of urine samples, are discussed. Pragmatic concepts summarized in this article should minimize the potential risks of misinterpreting urine drug screens.

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Urine drug screening: practical guide for clinicians.

Melanins are negatively charged, hydrophobic pigments of high molecular weight ([54][1], [88][2], [95][3], [139][4]) that are composed of polymerized phenolic and/or indolic compounds (Fig. [1][5]) ([45][6], [128][7]). Melanins are produced by organisms in all biological kingdoms, including a wide

http://www.reboundhealth.com/cms/images/pdf/virulencebacteriapigments%20id%2014023.pdf

Impact of melanin on microbial virulence and clinical resistance to antimicrobial compounds

Background: Methamphetamine (METH) and amphetamine (AMP) concentrations in 200 plasma and 590 oral fluid specimens were used to evaluate METH pharmacokinetics and pharmacodynamics after oral administration of sustained-release METH.

Methods: Eight participants received four oral 10-mg S -(+)-METH hydrochloride sustained-release tablets within 7 days. Three weeks later, five participants received four oral 20-mg doses. Blood samples were collected for up to 24 h and oral fluid for up to 72 h after drug administration.

Results: After the first oral dose, initial plasma METH detection was within 0.25–2 h; c max was 14.5–33.8 μg/L (10 mg) and 26.2–44.3 μg/L (20 mg) within 2–12 h. In oral fluid, METH was detected as early as 0.08–2 h; c max was 24.7–312.2 μg/L (10 mg) and 75.3–321.7 μg/L (20 mg) and occurred at 2–12 h. The median oral fluid-plasma METH concentration ratio was 2.0 across 24 h and was highly variable. Neutral cotton swab collection yielded significantly higher METH and AMP concentrations than citric acid candy-stimulated expectoration. Mean (SD) areas under the curve for AMP were 21% ± 25% and 24% ± 11% of those observed for METH in plasma and oral fluid, respectively. After a single low or high dose, plasma METH was >2.5 μg/L for up to 24 h in 9 of 12 individuals (mean, 7.3 ± 5.5 μg/L at 24 h); in oral fluid the detection window was at least 24 h (mean, 18.8 ± 18.0 μg/L at 24 h). The plasma and oral fluid 24-h METH detection rates were 54% and 60%, respectively. After four administrations, METH was measurable for 36–72 h (mean, 58.3 ± 14.5 h).

Conclusions: Perceived advantages of oral fluid for verifying METH exposure compared with urine include simpler specimen collection and reduced potential for adulteration, but urine offers higher analyte concentrations and a greater window of detection.

/pdf/methamphetamine-and-amphetamine-pharmacokinetics-in-oral-4goi4v2ghc.pdf

Methamphetamine and Amphetamine Pharmacokinetics in Oral Fluid and Plasma after Controlled Oral Methamphetamine Administration to Human Volunteers

Sweat testing for cocaine, codeine and metabolites by gas chromatography-mass spectrometry

Although the mechanism(s) of drug deposition in hair are unknown, there is evidence that suggests that the amount and type of melanin present are major factors in determining how much drug enters hair after exposure. The role of other hair components, such as lipids, has received less attention. We used in vitro binding techniques to evaluate the binding of radiolabeled cocaine to different types of treated and untreated hair specimens. Divided male and female Caucasoid (black/brown and blond colored) and Africoid (black colored) hair specimens (N = 7) were exhaustively extracted to remove lipid components (lipid-extracted hair). Separate portions were bleached to denature or alter melanin content. Experiments with radiolabeled cocaine were performed on untreated, lipid-extracted, and bleached portions of hair from different groups. Cocaine binding was significantly higher (p < .01) to male Africoid hair compared with other groups. The amount of drug binding was similar among female Africoid and male and female, black/brown Caucasoid specimens. The lowest amount of binding was observed with blond, female Caucasoid specimens. Binding experiments also revealed that specific cocaine binding generally did not differ significantly between lipid-extracted hair and untreated hair, but bleaching of most hair specimens resulted in significant (p < .01) decreases in specific binding compared with untreated hair. In separate experiments with cocaine-treated hair specimens, digested samples were evaluated to determine if removal of the insoluble melanin fraction from soluble hair components provided a means of normalization of drug content and elimination of color bias. Removal of the insoluble melanin fraction was not effective in removal of significant amounts of cocaine, which indicated that the digestion process released bound cocaine into the digest solution. Overall, these experiments suggested that lipids in hair play a minor role in drug binding, whereas melanin functions as a major binding site for cocaine. Natural (ethnic) or artificial (bleaching) differences in melanin content may determine the extent of cocaine entrapment in hair after drug exposure. Further, digestion of hair samples and removal of insoluble melanin failed to be effective in removal of hair color bias.

/pdf/in-vitro-binding-studies-of-drugs-to-hair-influence-of-2mlgu4fnsh.pdf

In Vitro Binding Studies of Drugs to Hair: Influence of Melanin and Lipids on Cocaine Binding to Caucasoid and Africoid Hair

Xenobiotics circulating in the blood may become incorporated into growing hair. Melanin has affinity for many pharmacologically unrelated drugs and is responsible for the pigmentation in hair. To assess the role of pigmentation in the incorporation of drugs into hair, the distribution of codeine and its metabolites was studied in Sprague-Dawley (SD; white nonpigmented hair), Dark Agouti (DA; brown pigmented hair), and hooded Long-Evans (LE; both black pigmented and white nonpigmented hair) rats. Codeine was administered at a dose of 40 mg/kg/day i.p. for 5 days. Fourteen days after beginning the dosing protocol, hair was collected and analyzed for codeine, and its metabolite, morphine, by positive-ion chemical ionization GC/ion-trap MS. The plasma pharmacokinetics for codeine and morphine were also determined after a single 40 mg/kg injection (equivalent to first dose in 5-day dosing protocol) in all three strains of rats. Hair and plasma codeine and morphine concentrations were also determined after acid hydrolysis to evaluate the presence of glucuronide metabolites. Codeine concentrations in the hair of SD, DA, and pigmented LE hair were 0.98 +/- 0.10, 5.99 +/- 1.24, and 111.93 +/- 18.69 ng/mg hair, respectively; morphine concentrations were 0.34 +/- 0.04, 0.51 +/- 0.11, and 14.46 +/- 1.81 ng/mg hair, respectively; morphine glucuronide concentrations were 0.67 +/- 0.08, 1.04 +/- 0.37, and 13.80 +/- 3.60 ng/mg hair, respectively. Studies examining the in vitro binding of [3H] codeine and [3H]morphine to hair demonstrated both specific and nonspecific binding sites for codeine and morphine. Pigmented hair from LE rats possessed the greatest number of binding sites, white hair from SD rats contained the least, and brown hair from DA rats was intermediate. A time course study of codeine and its metabolites showed pigment-mediated differences in incorporation of codeine and metabolites within a few hours of drug administration. These data indicate that pigmented hair possesses a greater capacity to bind and incorporate codeine and its metabolites than does nonpigmented hair. Interpretation of hair concentrations of drugs should involve consideration of hair pigmentation.

Incorporation of codeine and metabolites into hair. Role of pigmentation.

Allegations of illicit hydrocodone use have been made against individuals who were taking physician-prescribed oral codeine but denied hydrocodone use. Drug detection was based on positive urine opiate immunoassay results with subsequent confirmation of hydrocodone by gas chromatography-mass spectrometry (GC-MS). In these cases, low concentrations of hydrocodone (approximately 100 ng/mL) were detected in urine specimens containing high concentrations of codeine (> 5000 ng/mL). Although hydrocodone has been reported to be a minor metabolite of codeine in humans, there has been little study of this unusual metabolic pathway. We investigated the occurrence of hydrocodone excretion in urine specimens of subjects who were administered codeine. In a controlled study, two African-American and three Caucasian male subjects were orally administered 60 mg/70 kg/day and 120 mg/70 kg/day of codeine sulfate on separate days. Urine specimens were collected prior to and for approximately 30-40 h following drug administration. In a second case study, a postoperative patient self-administered 960 mg/day (240 mg four times per day) of physician-prescribed oral codeine phosphate, and urine specimens were collected on the third day of the dosing regimen. Samples from both studies were extracted on copolymeric solid-phase columns and analyzed by GC-MS. In the controlled study, codeine was detected in the first post-drug-administration specimen from all subjects. Peak concentrations appeared at 2-5 h and ranged from 1475 to 61,695 ng/mL. Codeine was detected at concentrations above the 10-ng/mL limit of quantitation for the assay throughout the 40-h collection period. Hydrocodone was initially detected at 6-11 h following codeine administration and peaked at 10-18 h (32-135 ng/mL). Detection times for hydrocodone following oral codeine administration ranged from 6 h to the end of the collection period. Confirmation of hydrocodone in a urine specimen was always accompanied by codeine detection. Codeine and hydrocodone were detected in all specimens collected from the postoperative patient, and concentrations ranged from 2099 to 4020 and 47 to 129 ng/mL, respectively. Analyses of the codeine formulations administered to subjects revealed no hydrocodone present at the limit of detection of the assay (10 ng/mL). These data confirm that hydrocodone can be produced as a minor metabolite of codeine in humans and may be excreted in urine at concentrations as high as 11% of parent drug concentration. Consequently, the detection of minor amounts of hydrocodone in urine containing high concentrations of codeine should not be interpreted as evidence of hydrocodone abuse.

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Identification of hydrocodone in human urine following controlled codeine administration

This study investigated the disposition patterns of cocaine and opiates into hair and fingernail specimens collected from 8 volunteers enrolled in a 10-week inpatient clinical study. All subjects were African-American males with a confirmed drug use history. Scalp hair and fingernail scrapings were collected weekly throughout the course of the study. Head hair was collected from the posterior vertex region, and fingernail scrapings were collected along the entire ventral surface of the nail plate. The specimens were introduced to successive decontamination washes including an isopropanol wash and three phosphate buffer washes. All decontamination washes were collected and analyzed. All specimens were enzymatically digested prior to being subjected to solid-phase extraction and derivatization. Analyses were performed using electron impact gas chromatography-mass spectrometry. Analytes investigated included eight cocaine analytes and five codeine analytes. The limit of quantitation for all analytes ranged from 0.1 to 0.5 ng/mg for both matrices. Cocaine was present at the highest concentrations of any analyte in both hair and nail. Benzoylecgonine and ecgonine methyl ester were the primary metabolites in both matrices and were typically less than 15% of cocaine concentrations. Codeine was the only opiate analyte identified in either hair or nail. Observed drug disposition profiles were different for hair and nails. A significant dose-response relationship was observed for hair specimens. The mean peak concentrations in hair after low dosing were half the concentration observed after high-dose administration. Generally, no clear relationship was evident between nail drug concentrations and dose. Decontamination washes removed less than 20% of the total drug present in hair, but removed most of the drug concentrations (60-100%) in nail. This investigation demonstrated that higher concentrations of drug were found in the subjects' hair than in their fingernails and that cocaine was found in both matrices at a greater concentration than codeine. Although both hair and nail have similar physical and chemical properties and may share common mechanisms of drug incorporation, this clinical study suggests that there are distinct differences in their disposition profiles.

/pdf/the-disposition-of-cocaine-and-opiate-analytes-in-hair-and-30epil8q3h.pdf

Robert E. Joseph

Papers

Sweat testing for cocaine, codeine and metabolites by gas chromatography-mass spectrometry

In Vitro Binding Studies of Drugs to Hair: Influence of Melanin and Lipids on Cocaine Binding to Caucasoid and Africoid Hair

Incorporation of codeine and metabolites into hair. Role of pigmentation.

Identification of hydrocodone in human urine following controlled codeine administration

The disposition of cocaine and opiate analytes in hair and fingernails of humans following cocaine and codeine administration